Melanoma is the most deadly skin cancer with an increasing incidence [1]. Over the past decade, it has been of interest to develop receptor-targeting radiolabeled peptides for melanoma imaging since early diagnosis followed by prompt surgical removal is a patient's best opportunity for a cure. Due to the over-expression on melanoma, both melanocotin-1 (MC1) and αvβ3 integrin receptors have been used as targets for radiolabeled alpha-melanocyte stimulating hormone (α-MSH) [2-14] and Arg-Gly-Asp (RGD) peptides [15-22], respectively. Recently, the inventors have developed a novel RGD-conjugated α-MSH hybrid peptide targeting both MC1 and αvβ3 integrin receptors for M21 human melanoma imaging [23]. The cyclic RGD {Arg-Gly-Asp-DTyr-Asp} motif was conjugated to [Cys3,4,10, D-Phe7, Arg11]α-MSH3-13} peptide via a lysine linker to yield RGD-Lys-(Arg11)CCMSH peptide. Meanwhile, the inventors designed two control peptides namely RAD-Lys-(Arg11)CCMSH and RGD-Lys-(Arg11)CCMSH scramble for comparison. The in vitro results revealed that the switch from RGD to RAD in the hybrid peptide decreased the αvβ3 integrin receptor binding affinity by 248-fold, whereas the scramble of CCMSH moiety in the hybrid peptide sacrificed the MC1 receptor binding affinity by 100-fold. The biodistribution results demonstrated that targeting both MC1 and αvβ3 integrin receptors enhanced the melanoma uptake of 99mTc-RGD-Lys-(Arg11)CCMSH in M21 human melanoma xenografts. The xenografted M21 human melanoma lesions were clearly visualized using 99mTc-RGD-Lys-(Arg11)CCMSH as an imaging probe [23].
While the switch from RGD to RAD in the hybrid peptide decreased the αvβ3 integrin receptor binding affinity by 248-fold, surprisingly, the inventors found that the switch from RGD to RAD in the hybrid peptide dramatically increased the MC1 receptor binding affinity of RAD-Lys-(Arg11)CCMSH compared to RGD-Lys-(Arg11)CCMSH (0.3 vs. 2.0 nM) in M21 melanoma cells [23]. Therefore, the inventors were interested in investigating whether such change in MC1 receptor binding affinity could result in enhanced melanoma uptake of 99mTc-RAD-Lys-(Arg11)CCMSH compared to 99mTc-RGD-Lys-(Arg11)CCMSH. Thus, the inventors examined the receptor binding affinity of RAD-Lys-(Arg11)CCMSH, internalization and efflux properties of 99mTc-RAD-Lys-(Arg11)CCMSH in B16/F1 melanoma cells. Furthermore, the inventors determined the melanoma targeting and imaging properties of 99mTc-RAD-Lys-(Arg11)CCMSH in B16/F1 melanoma-bearing C57 mice. The results proved to be unexpectedly favorable to prior art compounds.